Understanding the regulatory mechanisms by which voltage-dependent calcium channels (VDCCs) and their Ca2+ signals are regulated is central to the development of a mechanistic picture of nervous system function. The dynamics (or "shape") of intracellular Ca2+ signals help to specify which Ca2+-dependent effectors are triggered by a particular physiological stimulus. G protein-coupled receptors (GPCRs) provide one primary means of regulating VDCCs and the Ca2+ signals they generate; such regulation involves multiple mechanisms that converge on individual VDCCs to modify their gating, thereby generating unique patterns of Ca2+ influx. Despite their importance for nervous system function, however, the molecular mechanisms that control the rates of onset and termination of such GPCR-mediated modulation are not fully understood. In the process of identifying the molecules that determine the kinetics of Cav2.2 channel modulation by GPCRs, we have uncovered a novel mechanism for regulation Ca2+ influx through Cav2.2 channels that involves a receptor-dependent removal of the VDCC from the plasma membrane in dorsal root ganglion neurons. The GPCR-mediated internalization underlies in part the voltage-independent inhibitory component of the current. This is the first demonstration that GPCRs can alter the number of VDCCs at the plasma membrane. We've shown that Cav2.2 channels are associated with beta- arrestin1 and that this interaction is required for internalization. These data support the following hypothesis to be tested by experiments outlined in this proposal: Arrestin controls the timing of Ca2+ influx through Cav2.2 channels via direct interaction with the VDCC by a mechanism that is independent of its conventional role in regulating receptor-G protein coupling. As dorsal root ganglion neurons are central to normal pain transmission and neuropathic pain syndromes, results from our experiments have potential therapeutic applications in the development of new analgesics.